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• W3090757480 abstract "Abstract We present an optimized protocol for enhanced amplification and enrichment of viral DNA for Next Generation Sequencing of begomovirus genomes. The rapid ability of these viruses to evolve threatens many crops and underscores the importance of using next generation sequencing efficiently to detect and understand the diversity of these viruses. We combined enhanced rolling circle amplification (RCA) with EquiPhi29 polymerase and size selection to generate a cost-effective, short-read sequencing method. This optimized protocol produced short-read sequencing with at least 50% of the reads mapping to the viral reference genome. We provide other insights into common misconceptions about RCA and lessons we have learned from sequencing single-stranded DNA viruses. Our protocol can be used to examine viral DNA as it moves through the entire pathosystem from host to vector, providing valuable information for viral DNA population studies, and would likely work well with other CRESS DNA viruses. Highlights Protocol for short-read, high throughput sequencing of single-stranded DNA viruses using random primers Comparison of the sequencing of total DNA versus size-selected DNA Comparison of phi29 and Equiphi29 DNA polymerases for rolling circle amplification of viral single-stranded DNA genomes" @default.
• W3090757480 created "2020-10-08" @default.
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• W3090757480 date "2020-10-01" @default.
• W3090757480 modified "2023-09-26" @default.
• W3090757480 title "An improved experimental pipeline for preparing circular ssDNA viruses for next-generation sequencing" @default.
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• W3090757480 doi "https://doi.org/10.1101/2020.09.30.321224" @default.
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